Development of excluded ELISA and its application in the detection of HBsAg in sera with hepatitis B virus infection

Objective:To establish a simple two-step labeled immune test to detect the serum HBsAg.Methods:The excluded ELISA was established based on the previous G6 ELISA for serum HBsAg detection,and its relative color strength,sensitivity,and the antagonistic capacity to the Hooks effect were assessed using serial dilutions test.The assay was used to detect a group of clinical specimens,and the results were compared with those of the classic two-step and one-step ELISA.Results:The test signal strength of Excluded ELISA was roughly the same of the other two method,two-step and one-step ELISA in similar experimental conditions.The sensitivity was 19.2 pg/ml,12.6 pg/ml and 14.5 pg/ml,respectively.When HBsAg concentration of the samples was up to 37.5 μg/ml,one-step ELISA began to show the Hooks effect,and the measured values were close to the threshold value when HBsAg up to 2 400 μg/ml;But the excluded ELISA and the classical two-step ELISA both were showed similar antagonistic to the Hooks effect.By use of the three methods to detect 231 clinical samples,the results showed that there were 17 samples with moderate to severe Hooks effects when detected by one-step ELISA,acconunting for 15.2% in 112 samples with high-titer positive samples(confirmed by dilution experiments),whereas there was no Hooks effect detected by the other two methods.Conclusion:In the premise of guaranteed results,the excluded ELISA is equal to the one-step ELISA in the degree of simplification of operation,but it could completely overcome the interference of Hooks effect.